The invention relates to separation apparatus and more specifically to separation columns and methods for manufacturing separation columns for use in separation processes.
Separation-based analytical methods, including chromatography, electrophoresis and electrochromatography are useful in determining individual samples in complex mixtures. In chromatography, a sample to be analyzed is introduced into a separation column, which contains a mobile phase and a stationary phase. Components of the sample separate as the sample passes through the column due to differences in interaction of the different components with the stationary phase.
Electrophoresis is a separation technique that is based on the mobility of ions in an electric field. In capillary electrophoresis, a sample is placed in a capillary tube, which contains an electrophoretic medium. Upon application of an electric field across the capillary, components of the sample migrate at different rates towards the oppositely charged ends of the capillary based on their relative electrophoretic mobilities in the medium. Electrochromatography is a combination of chromatography and electrophoresis, in which the mobile phase is transported through the separation system by electroosmotic flow (EOF).
Separation of samples in complex mixtures based on analytical systems that are capable of executing large numbers of separations would be useful. In particular, separation technologies that process multiple samples quickly and multi-dimensional separations for each sample are desired. However, existing separation technologies do not generally have these features. Liquid chromatography systems do not readily adapt to parallel processing because adding multiple precision pumps and valves, necessary to deliver multiple samples, is both impractical and expensive. Multi-dimensional chromatography separations are feasible by transferring components from a first separation column to a second separation column with rotary valves. However, such processes can be very slow. Parallel processing for capillary electrophoresis has been achieved using microfabrication, which allows multiple channels to be placed on a single chip. However, a limitation is that no methods are available to introduce a large number of samples into the channels and to rapidly clean the sample metering channels between separations.
The present invention relates to separation columns for use in chromatography, electrochromatography and electrophoresis, which overcome the sample limitations of the existing separation systems. The separation columns of the present invention also require orders of magnitude less solvent and analyte, thereby diminishing the sample disposal problem.
In one aspect, the invention relates to a separation column, which is used in a separation process. The separation column includes multiple collocated monolith support structures and interconnected channels defined by the support structures. The collocated monolith support structures are arrayed in two dimensions to define channels that periodically split and merge. In one embodiment, the support structures are in communication with each other at the first end of each support structure and a cover plate is in communication with the support structures at the second end of each structure.
In another aspect, the invention relates to a method for manufacturing a separation column, which is used in a separation process. According to the method, a substrate is patterned to designate the areas of the substrate to be etched. The patterned substrate is etched to create multiple collocated monolith support structures arrayed in two dimensions and interconnected channels defined by the support structures. In one embodiment, a cover plate is attached on a surface of the created support structures to enclose the separation column. In another embodiment, the substrate is etched to create the support structures and the channels that are substantially uniform in shape and size. In yet another embodiment, the substrate is etched to create interconnected channels, in which each channel has an aspect ratio of from about 5 to about 100. The aspect ratio as used herein is the ratio of the depth to the width of a channel between adjacent monolith structures, where the depth is a dimension perpendicular to the surface of the substrate and the width is a dimension parallel to the surface of the substrate and perpendicular to the flow direction in the channel.
In another aspect, the invention relates to a separation column. The separation column includes an inlet, a separation region and an outlet. The separation region is in communication with the inlet and comprises a plurality of collocated monolith support structures that are arranged in two dimensions. The support structures define a plurality of interconnected channels that sequentially split and merge. The outlet is in communication with the separation region.
In yet another aspect, the invention relates to a separation apparatus. The separation apparatus includes a separation column, a plurality of reservoirs for mobile phases or buffers and a sample reservoir. The separation column has multiple collocated support structures arrayed in two dimensions and interconnected channels defined by the support structures. The reservoirs are in communication with the separation column. The sample reservoir is in communication with the separation column. In one embodiment, the separation apparatus also includes a pump for pumping a mobile phase from a reservoir through the separation column. In another embodiment, the separation apparatus also includes an electrophoresis apparatus.
In still another aspect, the invention relates to a method for separating components of a sample. According to the method, a medium solution is introduced into a separation column, which includes multiple collocated monolith support structures and interconnected channels defined by the support structures. A sample to be analyzed is also introduced into the separation column. The solution and the sample pass through the separation column in multiple streams and the multiple streams periodically intercouple. Components of the sample are separated as the sample passes through the column. The components may be separated by electrophoretic mobility, electroosmotic flow (EOF), EOF and partitioning with a stationary phase, micellar electrokinetic chromatography, or a combination of these.